This project will define and characterize the genes governing bacterial cell surface components of Rhizobium essential for the formation of nitrogen-fixing nodules in the roots of leguminous plants, and investigate aspects of physiology related to nodulation. To obtain nodulation-deficient (nod-) mutants with surface alterations will first be isolated by strong selective methods, and the nod- among them will be characterized with the genetic techniques developed previously by us and others. Selective methods include resistance to phage, ability to pass through Millipore filters, alteration in surface charge, and failure to bind specific antibody. The antibody method is a novel combination of Rhizobium genetics with the powerful new hybridoma technique for generating monoclonal antibodies with predetermined specificity, which will be used to disect genetically complex bacterial cell surface. This combination of hybridoma technology with bacterial genetics should have very wide applicability to the analysis of surface structure in other unrelated bacterial systems, including those of medical importance. Physiological studies will elucidate pathways of carbohydrate and energy metabolism, and investigate the role of nucleotide effectors in gene expression and nodulation.